The epithelia are extraordinarily diverse with respect to cell morphology, tissue structure, differentiation-related proteins expressed, and modes of growth regulation, befitting the wide variety of functions they perform. Much has been learned from cell culture experiments about the regulation of growth and differentiation in keratinocytes and mammary epithelial cells, but little is known about the biology of other epithelial cell types. Two epithelial cell types of embryonic mesodermal origin that we have cultured from normal human tissue have proven to be particularly interesting and amenable to the study of the regulation of keratin and vimentin expression in relation to their state of growth and differentiation. During the previous period of support we identified the in vitro culture requirements of normal human mesothelial cells. As a result, we found that this cell type is subject to reversible dedifferentiation as a consequence of rapid growth and loss of cell-cell contact. This dedifferentiation is manifest as a switching from an epithelioid morphology and predominantly keratin synthesis and content, to a fibroblastoid morphology and predominantly vimentin synthesis and content. The cells turn on keratin synthesis again and redifferentiate when their growth is arrested by growth factor deprivation and high cell density. We propose to use time-lapse video recording, indirect immunofluorescence using intermediate filament- and cell surface protein-specific antibodies, and in situ DNA-mRNA hybridization using keratin- and vimentin-specific cDNA sequences, to understand the role of growth rate, cell shape, and contact on mesothelial cell differentiation. We have recently found a nephron epithelial cell from the kidney cortex that undergoes a similar, but more gradual, dedifferentiation to a keratin-poor state which is not substantially reversed by growth arrest. We will use the above techniques to study the dedifferentiation process and use collagen gel culture and nude mouse transplantation to identify the extrinsic signals that promote differentiation in kidney epithelial cells and in embryonic nephrogenic mesoderm cells. Finally, we will study the relation between keratin/vimentin expression, steroid secretion, and ACTH sensitivity in adrenal cortex epithelial cells, which are also derived from embryonic mesoderm. These experiments will yield new information about the function of normal human mesothelial, kidney, and adrenal cells, thus providing a basis for using cell culture models to study age-, disease-, and developmentally-related changes in these tissues.